红掌细菌性叶疫病胁迫下实时荧光定量PCR(qRT-PCR)内参基因的筛选  

Reference Gene Selection for Quantitative Real-time PCR(q RT-PCR) in Anthurium(Anthurium andraeanum Linden) Tissues Under Bacterial Blight Stress

作　　者：洪阳[1,2,3] 尹俊梅[2,3] 黄少华[2,3] 李崇晖[2,3] 陆顺教[2,3] 杨澜[4] 牛俊海[2,3] 

机构地区：[1]海南大学农学院,海口570228 [2]中国热带农业科学院热带作物品种资源研究所/农业部华南作物基因资源与种质创制重点实验室,儋州571737 [3]海南省热带观赏植物种质创新利用工程技术研究中心,儋州571737 [4]贵州省园艺研究所,贵阳550006

出　　处：《农业生物技术学报》2015年第9期1178-1187,共10页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金青年项目(No.31201652);国家自然科学基金面上项目(No.31471902);海南省自然科学基金项目(No.314130);中央级公益性科研院所基本科研业务费专项资金(No.1630032012017)

摘　　要：地毯草黄单胞菌花叶万年青致病变种(Xanthomonas axonopodis pv.dieffenbachiae,Xad)引起的细菌性叶疫病是红掌(Anthurium andraeanum Linden)生产中最主要的病害。筛选出稳定可靠的内参基因,对于建立红掌叶疫病响应相关基因表达分析的实时荧光定量PCR(quantitative Real-time PCR,q RT-PCR)技术体系、研究红掌-病菌互作的分子病理机制具有意义。本研究选择6个常用候选基因包括延伸因子1α(elongation factor1-α,EF1α)、甘油醛三磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase,GADPH)、泛素7(ubiquitin 7,UBQ7)、组蛋白H3(histidine 3,His3)、α微管蛋白(α-tubulin,TUB)和β-肌动蛋白(β-Actin,ACTB),进行PCR扩增效率筛选,筛选红掌抗、感品种叶片和系统侵染进程中的表达稳定性。经ge Norm3.5、Best Keeper0.953及Normfinder1.0软件综合评价显示,不同候选基因的表达稳定性存在差异,其中UBQ7在红掌抗、感品种不同组织、时期表达均稳定,适合作为红掌叶疫病相关基因定量分析的内参基因,研究结果为红掌疫病应答的病理学研究提供了参考。Bacterial blight, caused by Xanthomonas axonopodis pv. dieffenbachiae （Xad）, is the most destructive disease in anthurium （Anthurium andraeanum Linden）. Appropriate reference gene selection is essential for accurate genes expression quantification about blight response by using quantitative Real-time PCR （qRT-PCR）, and to elucidate the phytopathology mechanism of anthurium-bacteria interaction. In this study, 6 commonly used candidates including elongation factorl-α （EF1α）, glyceraldehyde- 3- phosphate dehydrogenase （GADPI1）, ubiquitin 7（UBQ7）, histidine 3 （His3）, α-tubulin （TUB） and β-Actin （ACTB）, were employed to identify the most reliable reference genes. The genes amplification efficiency was assessed by qRT-PCR, and expression stability in the processes of foliage and systemic infection were analyzed on both resistant and susceptible cultivars, then generated dates were evaluated comprehensively using geNorm3.5, BestKeeper0.953 and NormFinder1.0 programs. As a result, the tested genes showed different expression characters in various samples, among which, the expression of UBQ7 was much more stable in different tissues and different varieties, could be recommended as reference gene for data normalization. The result will facilitate expression analysis of anthurium genes responding to blight.

关 键 词：红掌 细菌性叶疫病 实时荧光定量PCR(qRT-PCR) 内参基因 

分 类 号：S513[农业科学—作物学]